Why does adding SDS and potassium acetate, followed by

Last updated: 7/24/2022

Why does adding SDS and potassium acetate, followed by centrifugation precipitate the proteins and chromosomal DNA out of solution? Why does the RNA not precipitate out? This is for an RNA isolation & electrophoresis lab. Below is the explanation given in the lab manual: Detergents lyse membranes: SDS Cell membranes are composed of lipids and proteins rich in hydrophobic residues. SDS (sodium dodecyl sulfate) is a strong ionic detergent that solubilizes the plasma membrane. (Remember that in this protocol, E. coli, a prokaryote, is the source of RNA. Prokaryotes have cell walls which we need to break with lysozyme, but they do not have membrane bound organelles-meaning they do not have a nucleus-so we will need to separate the DNA from the RNA). In addition, SDS is also a strong protein denaturant as you learned when you isolated eukaryotic DNA in BIO130. In this protocol we use SDS plus potassium acetate to remove protein and other cellular constituents (discussed below under "Selective precipitation by SDS and potassium acetate"). Potassium acetate and SDS selectively precipitate protein and cell debris Sodium dodecyl sulphate is very soluble in water, while potassium dodecyl sulphate is very insoluble. In this procedure, we add potassium ions (potassium acetate), which form ionic bonds to produce insoluble potassium dodecyl sulphate. All of the molecules previously bound to SDS now precipitate from solution with the insoluble potassium dodecyl sulphate. By centrifuging this mixture, you remove most proteins, unbroken cells and fragments of cells, as well as most of the large, chromosomal DNA, which becomes trapped in the precipitate. Only very soluble molecules, such as RNA, small DNAs, and small salts remain in solution.